The human lens nucleus becomes progressively more yellow with age and with cataract formation. This nuclear color, measured accurately by the Fast Spectral Scanning Colorimetry (FSSC), a color measuring system, was found to affect strongly the contrast sensitivity function (CSF). The ability to correlate color parameters directly with visual dysfunction is a major accomplishment in vision research. However, the origin of nuclear color is still not definitely established. Three mechanisms can produce yellow pigments in in vitro experiments: photooxidation, nonenzymatic advanced glycation, and lipid peroxidation. It is hypothesized that lens pigments originate mostly from advanced glycation, since lens physiology favors the glycation mechanism. However, it is difficult to identify and quantify the glycation pigments or other pigments in the human lens by biochemical and biophysical measurements. This is because the pigment chromophores are labile to acid hydrolysis and they have similar molecular spectral features (UV-visible absorption and blue fluorescence). Immunochemical measurement may provide an alternative. In this proposal, the major objectives include: (i) to produce and characterize the pigments by in vitro advanced glycation, photooxidation of UV light, and lipid peroxidation; (2) to isolate fluorescent cross-linked species (pigmented peptides) and to use them as antigens in the preparation of polyclonal and monoclonal antibodies; and (3) to perform immunochemical measurements (Western blotting and ELISA) in an attempt to identify and quantify pigments in lens specimens from cataract surgery. This will permit us to define the predominant pigmentation mechanism that causes lens brunescence in a particular type of cataract. This knowledge may help in the development of a pharmaceutical means of slowing or stopping lens browning and could offer patients additional non-surgical options of treating cataract- and age-related vision loss.